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. 1990 Oct 1;271(1):59–66. doi: 10.1042/bj2710059

Role of cysteine residues in ribonuclease H from Escherichia coli. Site-directed mutagenesis and chemical modification.

S Kanaya 1, S Kimura 1, C Katsuda 1, M Ikehara 1
PMCID: PMC1150208  PMID: 2171503

Abstract

The role of the three cysteine residues at positions 13, 63 and 133 in Escherichia coli RNAase H, an enzyme that is sensitive to N-ethylmaleimide [Berkower, Leis & Hurwitz (1973) J. Biol. Chem. 248, 5914-5921], was examined by using both site-directed mutagenesis and chemical modification. Novel aspects that were found are as follows. First, none of the cysteine residues is required for activity. Secondly, chemical modification of either Cys-13 or Cys-133 with thiol-blocking reagents inactivates the enzyme, but that of Cys-63 does not. Thus the sensitivity of E. coli RNAase H to N-ethylmaleimide arises not from blocking of the thiol group but from steric hindrance by the modifying group incorporated at either Cys-13 or Cys-133.

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Selected References

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